Human AP (Allopregnanolone) ELISA Kit

Mfr.No.

EH2648

Sensitivity

0.094ng/ml

Detection Range

0.156-10ng/ml

Reactivity

Human

Assay Duration

2 hours

Assay Type

Competitive

Description

This kit is based on Competitive-ELISA detection method and takes 2h assay time. The microplate provided in this kit has been precoated with AP. AP in the sample or standard competes with a fixed amount of biotin-labeled AP for sites on a pre-coated antibody specific to AP. Free components are washed off. After adding HRP-Streptavidin Conjugate (SABC), biotin specifically binds with streptavidin to form immune complexes. Wash off unbound conjugate. Add TMB substrate solution. Then, TMB was catalyzed by HRP to produce a blue color product that turned yellow after adding a stop solution. Read the O.D. absorbance at 450nm in a microplate reader. Compare OD450 value of sample with the standard curve through curve fitting software to determine the concentration of AP in the sample. The concentration of the target substance is inversely proportional to the OD450 value.

Alternative Names

Allopregnanolone ELISA Kit, AP ELISA Kit

Detection Method

Coated with Antigen

Kit Components

Kit Components	Item	Size(48T)	Size(96T)	Storage Condition for Opened Kit
E001	ELISA Microplate(Dismountable)	8x6	8x12	Put the rest strips into a sealed foil bag with the desiccant. Stored for 1 month at 2-8°C; Stored for 6 month at -20°C
E002	Lyophilized Standard	1vial	2vial	Put the rest standards into a desiccant bag. Stored for 1 month at 2-8°C; Stored for 6 month at -20°C
E003	Biotin-labeled Antibody(Concentrated, 100X)	30ul	60ul	2-8°C (Avoid Direct Light)
E034	HRP-Streptavidin Conjugate(SABC, 100X)	60ul	120ul
E024	TMB Substrate	5ml	10ml
E039	Sample Dilution Buffer	10ml	20ml	2-8°C
E040	Antibody Dilution Buffer	5ml	10ml
E049	SABC Dilution Buffer	5ml	10ml
E026	Stop Solution	5ml	10ml
E038	Wash Buffer(Concentrated, 25X)	15ml	30ml
E006	Plate Sealer	3 pieces	5 pieces
E007	Product Description	1 copy	1 copy

Recovery

Add a certain amount of AP into the sample. Calculate the recovery by comparing the measured value with the expected amount of AP in the sample.

Sample Type	Recovery Range(%)	Average(%)
serum(n=10)	86-96	92
EDTA plasma(n=10)	91-104	97
Heparin plasma(n=10)	91-102	94

Linearity

Dilute the sample with a certain amount of AP at 1:2, 1:4 and 1:8 to get the recovery range.

Sample Type	1:2	1:4	1:8
serum(n=10)	85-103%	95-105%	86-104%
EDTA plasma(n=10)	82-96%	83-99%	89-99%
Heparin plasma(n=10)	81-98%	83-93%	81-100%

Specificity

Specifically binds with AP , no obvious cross reaction with other analogues.

Standard Curve

This product is detected by QC department and meets performance required in the manual. (Laboratory Humidity: 20%-60%; Temperature: 18°C -25°C; Equilibrate TMB substrate to 37°C before staining. After adding into the ELISA wells, incubate for 15min at 37°C in dark.) Due to different assay environments and operations, assay data below and standard curve are provided for reference. Experimenters should establish standard curve according to their own assay.

STD.(ng/ml)	OD-1	OD-2	Average
0	2.148	2.21	2.179
0.156	1.833	1.887	1.86
0.312	1.504	1.548	1.526
0.625	1.229	1.265	1.247
1.25	0.722	0.742	0.732
2.5	0.387	0.399	0.393
5	0.235	0.241	0.238
10	0.147	0.151	0.149

Components

Microplate,Lyophilized Standard,buffer,SABC,Label antibody,SABC,TMB,stop solution

Precision

Intra-assay Precision: samples with low, medium and high concentration are tested 20 times on the same plate.
Inter-assay Precision: samples with low, medium and high concentration are tested 20 times on three different plates.

Item	Intra-assay Precision			Inter-assay Precision
Sample	1	2	3	1	2	3
n	20	20	20	20	20	20
Mean (ng/ml)	0.29	1.3	5.11	0.31	1.26	5.08
Standard deviation	0.02	0.06	0.22	0.02	0.07	0.24
CV(%)	5.22	4.96	4.23	5.91	5.26	4.68

Background

AP/3α,5α-tetrahydroprogesterone/3α-hydroxy-5α-pregnan-20-one

Stability

Perform the stability test for the sealed kit at 37°C and 2-8°C and get relevant data.

ELISA kit(n=5)	37°C for 1 month	2-8°C for 6 months
Average(%)	80	95-100

Notes

For Research Use Only

Usage Notes

Step 1:
Take out the required plate wells, add 50ul Cap/Det Ab into each well, then add 50ul Standard or Sample into individual well. (When adding standard or sample, the disposable tip lightly touches the liquid level. Change the disposable tips for different samples and standards.) Gently tap the plate for 10s to ensure thorough mixing then static incubate for 60 minutes at 37°C.

Washing:
Wash the plate twice without immersion.

Step 2:
Add 100ul HRP-Streptavidin (orange) into each well, seal the plate and static incubate for 30 minutes at 37°C.

Washing:
Wash the plate five times without immersion.

Step 3:
Add 90ul TMB substrate solution, seal the plate and static incubate for 10-20 minutes at 37°C. (Accurate TMB visualization control is required.)

Step 4:
Add 50ul stop solution. Read at 450nm immediately and calculate.

Applications	ELISA
Applications Notes	AP ELISA Kit allows for the in vitro quantitative determination of AP concentrations in serum, plasma, tissue homogenates and other biological fluids.

Storage	2-8°C(Sealed), Don't cryopreserve.

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SKU	Supplier No.	Size	Your price	Quantity
BHE10802634	EH2648	48T	$280		Add to Cart
BHE10802634	EH2648	96T	$400		Add to Cart

Human AP (Allopregnanolone) ELISA Kit

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